Although ouabain-like molecules are widely distributed in the plant kingdom, their presence in the animal kingdom was thought to be limited to the large poison gland secretions of Bufonid toads. Based on the speculation that ouabain-like molecules would be useful regulators of (Na ion plus K ion)-ATPase of higher species, we examined the skin of a variety of amphibians for endogeneous ouabain activity, using a 3H-ouabain binding competition assay. Of the twenty species tested, 12 had ouabain-like activity demonstrable in skin, ranging from low levels to as high as 0.01% of the wet weight of skin. Activity was confirmed by ATPase and K ion flux inhibition studies. The activity is a small molecule, resistant to boiling and trypsin inactivation. It is suggested that some or all of this ouabain-like activity may play a role in the regulation of (Na ion plus K ion)-ATPase in amphibian skin.